The G-protein G13 but not G12 mediates signaling from lysophosphatidic acid receptor via epidermal growth factor receptor to Rho.
Lysophosphatidic acid (LPA) utilizes a G-protein-coupled receptor to activate the small GTP-binding protein Rho and to induce rapid remodeling of the actin cytoskeleton. We studied the signal transduction from LPA receptors to Rho activation. Analysis of the G-protein-coupling pattern of LPA receptors by labeling activated G-proteins with [alpha-32P]GTP azidoanilide revealed interaction with proteins of the Gq, Gi, and G12 subfamilies. We could show that in COS-7 cells, expression of GTPase-deficient mutants of Galpha12 and Galpha13 triggered Rho activation as measured by increased Rho-GTP levels. In Swiss 3T3 cells, incubation with LPA or microinjection of constitutively active mutants of Galpha12 and Galpha13 induced formation of actin stress fibers and assembly of focal adhesions in a Rho-dependent manner. Interestingly, the LPA-dependent cytoskeletal reorganization was suppressed by microinjected antibodies directed against Galpha13, whereas Galpha12-specific antibodies showed no inhibition. The tyrosine kinase inhibitor tyrphostin A 25 and the epidermal growth factor (EGF) receptor-specific tyrphostin AG 1478 completely blocked actin stress fiber formation caused by LPA or activated Galpha13 but not the effects of activated Galpha12. Also, expression of the dominant negative EGF receptor mutant EGFR-CD533 markedly prevented the LPA- and Galpha13-induced actin polymerization. Coexpression of EGFR-CD533 and activated Galpha13 in COS-7 cells resulted in decreased Rho-GTP levels compared with expression of activated Galpha13 alone. These data indicate that in Swiss 3T3 cells, G13 but not G12 is involved in the LPA-induced activation of Rho. Moreover, our results suggest an involvement of the EGF receptor in this pathway.
Gohla
A
A
Institut für Pharmakologie, Freie Universität Berlin, Thielallee 67-73, D-14195 Berlin, Germany.
Harhammer
R
R
Schultz
G
G
D016475:3T3 Cells; D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D008246:Lysophospholipids / Q000494:pharmacology; D051379:Mice; D009570:Nitriles / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled; D049368:Receptors, Lysophosphatidic Acid; D015398:Signal Transduction; D020032:Tyrphostins
D004791:Enzyme Inhibitors; D008246:Lysophospholipids; D009570:Nitriles; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D049368:Receptors, Lysophosphatidic Acid; D020032:Tyrphostins; C079836:tyrphostin 25; D066246:ErbB Receptors; D019204:GTP-Binding Proteins